Fig. 7.

SMAD6 is necessary for TGF-β₂-mediated inhibition of IL-1β-induced IL-8 and IL-6 secretion in H4 cells. To determine whether SMAD6 mediates the immunomodulatory effect of TGF-β₂ in H4 cells, the cells were transfected with siRNA oligonucleotide specific for SMAD6 or control siRNA. Acceptable transfection efficiency was achieved as assessed by detecting incorporation of Alexa Fluor into cells 24 h after transfection (a). SMAD6 siRNA resulted in 96% reduction in SMAD6 mRNA expression 48 h after transfection as compared to cells transfected with control siRNA (b). IL-1β induced a significant increase in both IL-8 (c) and IL-6 (d) secretion, and this was attenuated by TGF-β₂ in control cells. However, SMAD6 siRNA abolished the inhibition of IL-1β-induced production of IL-8 (c) and IL-6 (d) by TGF-β₂. These data suggest that SMAD6 is necessary for an anti-inflammatory effect of TGF-β₂. c ∗ p < 0.05 compared with IL-1β-induced secretion of IL-8 in control. d ∗ p < 0.05 compared with IL-1β-induced secretion of IL-6 in control.